Revision 9
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IHC-Bond, IHC-P, IF-IC, FC-FP

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

52-65

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q15116

Entrez-Gene Id:

5133

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:200
IHC Leica Bond 1:100 - 1:400
Immunohistochemistry (Paraffin) 1:100 - 1:400
Immunofluorescence (Immunocytochemistry) 1:200 - 1:400
Flow Cytometry (Fixed/Permeabilized) 1:100 - 1:200

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier free (BSA and azide free) version of this product see product #63815.

Specificity / Sensitivity

PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb recognizes endogenous levels of total PD-1 protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ala274 of human PD-1 protein.

Background

The programmed cell death 1 protein (PD-1, PDCD1, CD279) is a member of the CD28 family of immunoreceptors that regulate T cell activation and immune responses (1-3). The PD-1 protein contains an extracellular Ig V domain, a transmembrane domain, and a cytoplasmic tail that includes an immunoreceptor tyrosine-based inhibitory motif (ITIM) and an immunoreceptor tyrosine-based switch motif (ITSM). PD-1 is activated by the cell surface ligands PD-L1 and PD-L2 (4). Upon activation, PD-1 ITIM and ITSM phosphorylation leads to the recruitment of the protein tyrosine phosphatases SHP-1 and SHP-2, which suppress TCR signaling (5-7). In addition to activated T cells, PD-1 is expressed in activated B cells and monocytes, although its function in these cell types has not been fully characterized (8). The PD-1 pathway plays an important role in immune tolerance (3); however, research studies show that cancer cells often adopt this pathway to escape immune surveillance (9). Consequently, blockade of PD-1 and its ligands is proving to be a sound strategy for neoplastic intervention (10).

  1. Ishida, Y. et al. (1992) EMBO J 11, 3887-95.
  2. Shinohara, T. et al. (1994) Genomics 23, 704-6.
  3. Nishimura, H. et al. (1999) Immunity 11, 141-51.
  4. Freeman, G.J. et al. (2000) J Exp Med 192, 1027-34.
  5. Yokosuka, T. et al. (2012) J Exp Med 209, 1201-17.
  6. Sheppard, K.A. et al. (2004) FEBS Lett 574, 37-41.
  7. Chemnitz, J.M. et al. (2004) J Immunol 173, 945-54.
  8. Thibult, M.L. et al. (2013) Int Immunol 25, 129-37.
  9. Dong, H. et al. (2002) Nat Med 8, 793-800.
  10. Topalian, S.L. et al. (2012) Curr Opin Immunol 24, 207-12.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IHC-Bond: IHC Leica Bond IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 9
#86163

PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb

Western Blotting Image 1: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from human CD4+ T cells, MOLT-4, and Jurkat cells using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb (upper), and β-Actin (D6A8) Rabbit mAb #8457 (lower). CD4+ T cells were purified from human blood and stimulated for 9 days using beads coated with CD3 and CD28 antibodies in the presence of human interleukin-2 (hIL-2) #8907 (6.7 ng/ml).
Immunoprecipitation Image 1: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Immunoprecipitation of PD-1 protein from Molt-4 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb. Western blot analysis was performed using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb.
Immunohistochemistry Image 1: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human infiltrating papillary carcinoma of the breast using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb performed on the Leica® BOND Rx.
Immunohistochemistry Image 1: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon carcinoma using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb.
Immunohistochemistry Image 2: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human B-cell non-Hodgkin lymphoma using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb.
Immunohistochemistry Image 3: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded 293 cell pellets, control (left) or PD-1 transfected (right), using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb.
Immunohistochemistry Image 4: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Multiplex immunohistochemical analysis of paraffin-embedded human breast carcinoma usng PD-1 (Intracellular Domain) (D4W2J) XP® rabbit mAb (green), PD-L1 (E1L3N®) XP® rabbit mAb #13684 (red), LAG3 (D2G4O) XP® rabbit mAb #15372 (magenta), TIM-3 (D5D5R) XP® rabbit mAb #45208 (yellow), CD8α (C8/144B) mouse mAb #70306 (orange), and Pan-keratin (C11) mouse mAb #4545 (cyan).
Immunohistochemistry Image 5: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Multiplex immunohistochemical analysis of paraffin-embedded human ovarian carcinoma using PD-1 (Intracellular Domain) (D4W2J) XP® rabbit mAb (green), B7-H3 (D9M2L) XP® rabbit mAb #14058 (red), B7-H4 (D1M8I) XP® rabbit mAb (cyan), LAG3 (D2G4O) XP® rabbit mAb #15372 (magenta), TIM-3 (D5D5R) XP® rabbit mAb #45208 (yellow), and VISTA (D1L2G) XP® rabbit mAb #64953 (orange).
Immunofluorescence Image 1: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of MOLT-4 cells (left, positive) or HeLa cells (right, negative) using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb (green), Cytochrome c (6H2.B4) Mouse mAb #12963 (red), and DAPI #4083 (blue).
Flow Cytometry Image 1: PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb Expand Image
Flow cytometric analysis of fixed and permeabilized human peripheral blood mononuclear cells, untreated (left column) or treated with anti-CD3 (10ug/ml, 72hr) and anti-CD28 (5ug/ml, 72 hr; right column), using PD-1 (Intracellular Domain) (D4W2J) XP® Rabbit mAb #86163 (top row) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (bottom row), and co-stained with CD3 (UCHT1) Mouse mAb (APC Conjugate) #19881. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.